Carnosinase Gene—Is It Responsible for Diabetic Nephropathy?
Authors: Janssen B, Hohenadel D, Brinkkoetter P, Peters V, Rind N, Fischer C, Rychlik I, Cerna M, Romzova M, de Heer E, Baelde H, Bakker SJ, Zirie M, Rondeau E, Mathieson P, Saleem MA, Meyer J, Ko[Combining Diaeresis]ppel H, Sauerhoefer S, Bartram CR, Nawroth P, Hammes HP, Yard BA, Zschokke J, van der Woude FJ
Journal: Journal of the American Society of Nephrology (2005)

Despite hyperglycemia, not all diabetic patients develop nephropathy. The risk of developing nephropathy clusters within families, both in type 1 (1) and type 2 diabetes (2–5). Clustering of cardiovascular events and higher cardiovascular mortality have been observed in families with diabetes and nephropathy (6,7). Many genes have been considered as candidate genes for the development of nephropathy in diabetes: An association was observed between parental hypertension and nephropathy in offspring with type 1 diabetes (8–10) and higher BP were found in offspring of type 2 diabetic parents with nephropathy (5). Great interest had initially been focused on the D/I polymorphism of the angiotensin-converting enzyme (ACE) gene (11), based also on suggestive animal experiments (12), but a meta-analysis showed at best a modest correlation to the risk of nephropathy (13). Further interest had been generated by the sodium-lithium countertransport system (14), which predicts onset of diabetic nephropathy (15), but again, no evidence of genetic linkage had been forthcoming. More recent candidate genes were adiponectin (16), PPARγ (17) and others. Apart from looking at “candidate genes,” a strategy which is based on considerations of a priori plausibility and pathomechanistic hypotheses, another approach is to scan the genome for evidence that certain loci are associated with a given disease. In 2002 Vardarli et al. had reported that a locus on the long arm of chromosome 18 (18q22.3-23) was associated with higher susceptibility for diabetic nephropathy in 18 Turkish families (18), but the relevant gene had not been identified. Although other studies found, apart from several other loci, also a high logarithm of the odds (lod) score (an index of association) for this specific locus (19), the findings were not consistent in all studies (20). The study by Janssen et al. now showed, however, not only linkage to the locus on chromosome 18, but provided also plausible evidence for the involvement of one specific enzyme as well as a presumed pathomechanism explaining the link to diabetic nephropathy via two well-known culprits in the genesis of this condition, i.e., oxidative stress and advanced glycation end products (AGE). The authors identified carnosinase, i.e., the enzyme that degrades the dipeptide β-alanyl-L-histidine, as the culprit. Two isoforms of this enzyme exist, and they are coded for at this locus on chromosome 18 in a head-to-tail configuration: one isoenzyme is secreted and demonstrable in the circulation, it has 12 exons, and it is coded for by the gene CNDP1; the other is located intracellularly, it also has 12 exons, and it is coded for by the gene CNDP2. The authors screened 5500 diabetic patients and selected for genetic analysis 135 patients with long-lasting diabetes and diabetic nephropathy according to stringent criteria, and 107 diabetic patients without diabetic nephropathy. The first step was to look for gross truncating deletions by complete sequencing analysis in a limited number of patients, but none were found. In a second step, an exploratory analysis, they looked for associations with polymorphisms and identified a trinucleotide sequence in the 5′ region, i.e., exon 2 of CNDP1. This trinucleotide sequence, coding for leucine in the leader sequence, yielded a string of 5, 6, or 7 leucines. The allele with 5 leucines was more frequent in diabetic patients without nephropathy (88%) than in those with nephropathy (59%). In a further step, the authors investigated whether homozygosity for the apparently protective allele was more frequent in diabetic patients without nephropathy, and this was indeed the case: Homozygosity was found in 25 of 63 type 2 diabetic patients without diabetic nephropathy, but only in 20 of 77 diabetic patients with nephropathy, and the same was found in patients with type 1 diabetes. The next step was to measure serum carnosinase activity, and it was indeed lowest in patients homozygous for the carriers of the gene with 5 trinucleotides. The serum carnosine concentrations were all extremely low, but this may not reflect the local concentrations at target organs, specifically the kidney, where the enzyme is also expressed, particularly in podocytes as documented in orientating studies. The findings suggest the following sequence: Low carnosinase activity in carriers of the favorable low trinucleotide sequence, particularly in the homozygous constellation, causes high carnosine levels, and the latter are protective. If this is true, addition of carnosine to renal cells should prevent processes thought to be relevant in the genesis of diabetic nephropathy. To prove this point, the authors added carnosine to mesangial cells or podocytes incubated in the presence of a hyperglycemic medium, and indeed this reduced the expression of TGF-β2 by mesangial cells and of collagen IV and fibronectin by podocytes. The findings suggest that a higher number of trinucleotides acts functionally as a gain of function trait consistent with a dominant mode of inheritance. What is known about carnosine? It prevents cellular senescence (21) and is said to have anti-aging properties (22). It scavenges reactive oxygen species (23) and reacts with macromolecular carbonyls (22). It also inhibits ACE (24). These are all properties that would interfere with initiation and progression of diabetic nephropathy and conceivably also with other micro- and macrovasular complications of diabetes. We shall certainly hear about this interesting substance in the future.